CN-122011214-A - Duck NMB protein polyclonal antibody and preparation method and application thereof

Abstract

The invention discloses a duck NMB protein polyclonal antibody, a preparation method and application thereof, and belongs to the technical field of biology. The duck NMB protein polyclonal antibody is prepared by taking duck NMB fusion protein as an antigen and immunizing animals. The amino acid sequence of the duck NMB fusion protein is shown as SEQ ID NO. 6. The duck NMB fusion protein contains all antigenic sites of NMB protein, and the recombinant strain can efficiently express soluble fusion protein. The polyclonal antibody of the duck NMB protein prepared by taking the polyclonal antibody as an antigen has the titer of 1:51200, can specifically bind with NMB protein in duck tissues, can accurately detect the expression and distribution of the NMB protein by Western Blot and IHC, provides core support for researching and developing the duck NMB protein detection kit, has low overall technical cost and easy operation, and is suitable for the related research of the duck NMB protein and the development of detection products.

Inventors

• MA ZHIYU
• LI JIA
• CHEN GUANGZHEN
• ZHANG YING
• FAN JIANHUA
• PAN SHIFENG
• ZHANG CHENGCHENG
• HE ZHIPEI
• YANG SHENG
• WANG YIWEN

Assignees

• 扬州大学
• 江苏省家禽科学研究所

Dates

Publication Date

20260512

Application Date

20260310

Claims (10)

1. 1. The duck NMB fusion protein is characterized in that the amino acid sequence of the duck NMB fusion protein is shown as SEQ ID NO. 6.
2. 2. The duck NMB protein polyclonal antibody is characterized in that the duck NMB protein polyclonal antibody is prepared by taking the duck NMB fusion protein as an antigen and immunizing animals.
3. 3. The method for preparing the duck NMB protein polyclonal antibody according to claim 2, comprising the steps of: (1) Transferring a recombinant vector containing the encoding gene of the duck NMB fusion protein of claim 1 into a host cell, and performing induced expression and purification to prepare the duck NMB fusion protein; (2) And immunizing animals by taking the duck NMB fusion protein as an antigen, and collecting serum to obtain the duck NMB protein polyclonal antibody.
4. 4. The preparation method according to claim 3, wherein the encoding gene of the duck NMB fusion protein is shown in SEQ ID NO. 5.
5. 5. The method according to claim 3, wherein the conditions for inducing expression are such that IPTG at 25℃is used in an amount of 0.1-1.5nM for induction of 10-16 h.
6. 6. The method of claim 3, wherein the method of purification comprises a His-tag protein purification method.
7. 7. A method of preparation according to claim 3 wherein the animal is a rabbit.
8. 8. Use of the polyclonal antibody against duck NMB protein as claimed in claim 2 for the preparation of a product for detecting duck NMB protein.
9. 9. The use according to claim 8, wherein the product comprises a kit for detecting NMB protein expression in duck organs, tissues or cells.
10. 10. The use according to claim 9, wherein the kit comprises a Western Blot kit or an immunohistochemical kit.

Description

Duck NMB protein polyclonal antibody and preparation method and application thereof Technical Field The invention relates to the technical field of biology, in particular to a duck NMB protein polyclonal antibody, a preparation method and application thereof. Background Neuromedin B (Neuromedin B, NMB) is one of the bombesin (Bombesin, BN) families, widely expressed in various tissues and organs of the animal body, involved in regulating various biological functions of the body, such as regulating gastrointestinal motility and hormone secretion, affecting energy metabolism homeostasis, mediating stress and fear-related behaviors, maintaining normal tissue growth and involved in tumor development, regulating immunity and reproduction, etc. The Open Reading Frame (ORF) of the duck NMB gene is 387 bp, codes for 128 aa, and has high homology with chicken waterfowl NMB protein. In addition, the NMB gene of the duck has higher expression level in central and peripheral tissues and organs such as brain, cerebellum, optic leaf, trachea, lung, pineal gland, thyroid gland, kidney, tail fat gland and the like, which indicates that NMB possibly has important physiological functions in the duck body. As an important waterfowl, the duck not only has important ecological value, but also is an important economic and cultural resource for human beings. Therefore, ensuring the health of the ducks, improving the production performance of the ducks and preventing the occurrence and the transmission of duck diseases are important responsibilities of livestock and veterinary work. Meanwhile, the duck is taken as an important waterfowl, and the related research of the duck is developed to provide reference for the research of other waterfowl. Because of high cost and poor stability of artificially synthesized duck NMB protein, and depending on equipment and process. Therefore, the preparation of the soluble duck NMB fusion protein by using the prokaryotic expression system has remarkable advantages, can provide more abundant structural information, wider epitope coverage and stronger immunogenicity, so that the polyclonal antibody and the monoclonal antibody which are efficient and reliable in various immunological applications are more likely to be obtained, and the main risks and application limitations of a polypeptide method caused by simple structure and single epitope are effectively avoided. However, the prior art lacks soluble expression of duck NMB protein and related research on preparation of polyclonal antibody of duck NMB protein. Disclosure of Invention The invention aims to provide a duck NMB protein polyclonal antibody, a preparation method and application thereof, so as to solve the problems in the prior art. The duck NMB fusion protein contains all antigenic sites of NMB protein, and the recombinant strain can efficiently express soluble fusion protein. The polyclonal antibody of the duck NMB protein prepared by taking the polyclonal antibody as an antigen has the titer of 1:51200, can specifically bind with NMB protein in duck tissues, can accurately detect the expression and distribution of the NMB protein by Western Blot and IHC, provides core support for researching and developing the duck NMB protein detection kit, has low overall technical cost and easy operation, and is suitable for the related research of the duck NMB protein and the development of detection products. In order to achieve the above object, the present invention provides the following solutions: the invention provides a duck NMB fusion protein, and the amino acid sequence of the duck NMB fusion protein is shown as SEQ ID NO. 6. The invention also provides a duck NMB protein polyclonal antibody, which is prepared by taking the duck NMB fusion protein as an antigen and immunizing animals. The invention also provides a preparation method of the duck NMB protein polyclonal antibody, which comprises the following steps: (1) Transferring a recombinant vector containing the encoding gene of the duck NMB fusion protein into a host cell, and performing induced expression and purification to prepare the duck NMB fusion protein; (2) And immunizing animals by taking the duck NMB fusion protein as an antigen, and collecting serum to obtain the duck NMB protein polyclonal antibody. Further, the encoding gene of the duck NMB fusion protein is shown as SEQ ID NO. 5. Further, the conditions for inducing expression are such that 10-16 h is induced at 25℃using 0.1-1.5nM IPTG. Alternatively, the purification method comprises His-tag protein purification. Optionally, the animal is a rabbit. The invention also provides application of the duck NMB protein polyclonal antibody in preparation of products for detecting duck NMB protein. Further, the product comprises a kit for detecting NMB protein expression in duck organs, tissues or cells. Further, the kit comprises a Western Blot kit or an immunohistochemical kit. The invention discloses the following technical